Photodynamic plant defoliants

ABSTRACT

Plant defoliating compositions comprising δ-aminolevulinic acid and one or more chlorophyll biosynthesis modulators; and methods of making and using same.

The invention described herein was made in the course of work supportedby grants from the U.S. Department of Agriculture, the National ScienceFoundation, the University of Illinois, the University of IllinoisAgriculture Experiment Station and the John P. TrebellasPhotobiotechnology Research Endowment.

This application is a division of application Ser. No. 07/521,119, filedMay 3, 1990,now U.S. Pat. No. 5,163,990, issued Nov. 17, 1992, which isa continuation-in-part of application Ser. No. 06/895,529 filed Aug. 11,1986, now U.S. Pat. No. 5,127,938 issued Jul. 7, 1992, which is acontinuation of application Ser. No. 06/754,092 filed Jul. 15, 1985, nowabandoned, which in turn is a continuation-in-part of application Ser.No. 06/634,932 filed Jul. 27, 1984, now abandoned.

FIELD OF THE INVENTION

This invention pertains to plant desiccating compositions and methods,and more particularly to plant desiccating compositions and methods forthe induction of the accumulation of photodynamic tetrapyrroles in thefoliage of plants.

BRIEF DESCRIPTION OF THE FIGURES

The following terms, as used hereinbelow, have the following meaningunless expressly stated to the contrary: Alk=(C₁ -C₁₀)alkyl group;ALA=δ-aminolevulinic acid; Chl=chlorophyll; Chlide a=chlorophyllide a;coprogen=coproporphyrinogen; cv=cultivar; dicot=dicotyledenous plant;DP=dipyridyl; DV=divinyl; E=ester; F.A1=fatty alcohol; LWMP=longerwavelength metalloporphyrins (the putative intermediates of ring Eformation); M=methylation; ME =methyl ester; Me - methyl;Me.P=methylpropionate; monocot=monocotyledenous plant;MPE=Mg-protoporphyrin monoester; MP(E)=mixture of MPE andMg-protoporphyrin IX; MV=monovinyl; P=esterification with geranylgeraniol, followed by stepwise conversion of the latter to phytol;PBG=porphobilinogen; Pchl= protochlorophyll;Pchlide=protochlorophyllide; Phy=phytol; Proto=protoporphyrin IX;Protogen=protoporphyrinogen IX; Urogen=uroporphyrinogen, var=variety.

The invention will be understood more clearly and fully with referenceto the accompanying figures, in which:

FIG. i depicts the six-branched Chl a biosynthetic pathway;

FIG. 2 depicts representative structures of some of themetallotetrapyrroles ("tetrapyrroles") depicted in FIG. 1;

FIG. 3 depicts the percent defoliation of apple seedlings as related tothe accumulation of protoporphyrin IX (^(*) coefficient of determination(r²) is significant at the 5% level);

FIG. 4 depicts the percent defoliation of apple seedlings as related tothe accumulation of divinyl Mg protoporphyrin monoester (^(*)coefficient of determination (r²) is significant at the 5% level); and

FIG. 5 depicts the percent defoliation of apple seedlings as related tothe accumulation of monovinyl protochlorophyllide (^(*) coefficient ofdetermination (r²) is significant at the 5% level).

BACKGROUND OF THE INVENTION

The timing and manipulation of plant development have some importantimplications in deciduous tree and crop production. Two importantaspects in the manipulation of plant development are defoliation andfruit drop. First, controlled defoliation of nursery stock is essentialfor the effective management of rootstocks and grafted trees of bothfruit and woody ornamental crops. For example, in areas with longgrowing seasons, nurserymen need to hasten defoliation in order tofacilitate autumn digging. Autumn digging is a process whereby trees aredug out of the ground or "undercut" and then placed in cold storage. Inorder for a tree to be undercut and placed in cold storage, it must bedormant, i.e., it is not producing new shoots. Defoliation triggersdormancy in the tree which is followed by hardening of the tree.Hardening protects the tree from cold injury. Young apple trees areusually undercut and removed from the nursery in the fall and placed incold storage. However, very often, trees in the nursery become juvenileby continuing to produce new shoots and retaining foliage longer throughthe season which retards hardening of the trees, making them moresusceptible to cold injury. Therefore, it is desirable to hastendefoliation of the young trees.

Prior to the advent of chemical sprays or dust treatments, defoliationwas done by hand, sometimes causing damage to shoots, bark, and buds.However, hand defoliation is time consuming and adds to the cost of anursery operation. Furthermore, nutrients that would normally betranslocated into the shoots during leaf senescence are lost.

Second, control of fruit drop is important in harvesting of fruit fromtrees. For example, aerial and subterranean fruits and vegetables arepresently harvested either manually or mechanically. Manual harvestingis labor-intensive and expensive. Aerial mechanical harvesting of fruitsuses heavy equipment that shakes the fruit off the tree. This in turncauses both soil compaction and frequent limb and trunk injury resultingin shorter tree life. Such mechanical harvesting also results in fruitdamage in the form of bruises, cuts and punctures.

Various naturally occurring and non-naturally occurring chemicalsubstances have been used for the purposes of defoliation andenhancement of fruit drop. In the case of fruit drop, chemicals are usedto reduce fruit-pedicel attachment strength, thus allowing tree shakersto drop fruit more easily. However, chemical fruit harvesting has notbeen completely successful and has been used only in order to facilitatemechanical harvesting.

It would be a significant and useful advance in the art to have achemical composition capable of causing defoliation and/or defoliationand fruit drop in plants, particularly deciduous fruit trees, via amechanism involving one or more naturally occurring intermediates in thechlorophyll biosynthesis and which alleviates the disadvantagesassociated with present methods for defoliation and fruit drop.

Chlorophyll biosynthesis is a major biological phenomenon in thebiosphere and is mandatory for the biosynthesis of photosyntheticmembranes during greening and for the repair and maintenance of the Chlin mature green plants. The chlorophylis are a group of Mg-tetrapyrroleswhich in green plants catalyze the conversion of solar energy intochemical energy via the process of photosynthesis. There are two basicclasses of chlorophyll, designated chlorophyll a (Chl a) and chlorophyllb (Chl b); Chl a is involved in the collection of solar energy and itsconversion to chemical energy whereas Chl b is believed to be involvedonly in the collection of solar energy.

As shown in FIG. 1, ten species of Chl a are all synthesized via amultiple-branched pathway from one common precursor, δ-aminolevulinicacid (ALA), via a series of porphyrin, Mg-porphyrin, andprotochlorophyll intermediates, collectively referred to astetrapyrroles or tetrapyrrole intermediates (see FIG. 2).

As can be seen in FIG. 1, three of the branches of the synthetic pathwayhave been designated as divinyl (DV) pathways; the two monocarboxylicacid pathways predominate in dicots and in monocots in the presence oflight. The remaining three branches have been designated the monovinyl(MV) pathways; the two monocarboxylic acid pathways predominate inmonocots in the dark. Plants may be classified as "monovinyl" or"divinyl" plants, depending on .which pathways predominate. A monovinylplant is a plant species which in darkness accumulates MV Pchlide viathe MV monocarboxylic acid biosynthetic routes and upon exposure tolight initially forms Chl mainly via the MV monocarboxylic acid routes.Divinyl plants are plant species which accumulate mainly DV Pchlide indarkness and upon exposure to light initially form Chl preferably viathe DV monocarboxylic biosynthetic routes. After several hours indaylight both MV and DV plants appear to form Chl via the DVmonocarboxylic routes. This in turn has led to the classification ofplants into four different greening groups (Rebeiz, C. A.,Montazer-Zouhoor, A., Mayasich, J. M., Tripathy, B. C., Wu, S., andRebeiz, C. C. CRC Crit. Rev. in Plant Sci., 6:385-435 (1988)):

(a) Dark divinyl/light divinyl (DDV/LDV). In this greening group,chlorophyll formation proceeds via the DV-enriched protochlorophyllidepools at daybreak and in daylight.

(b) Dark monovinyl/light divinyl (DMV/LDV). In this greening group,chlorophyll formation proceeds via the MV-enriched protochlorophyllidepools at daybreak and via the DV-enriched protochlorophyllide pools indaylight.

(c) Dark monovinyl/light monoVinyl (DMV/LMV). In this greening group,chlorophyll formation proceeds via the MV-enriched protochlorophyllidepools in darkness and via the MV-enriched protochlorophyllide pools atdaybreak and in daylight.

(d) Dark divinyl/light monovinyl (DDV/LMV). In this pathologicalgreening group, chlorophyll formation proceeds via the DV-enrichedprotochlorophyllide pools at daybreak and via the MV-enrichedprotochlorophyllide pools in daylight.

As can be seen from FIG. 2, δ-aminolevulinic acid (ALA) is a 5-carbonamino acid. ALA is found in most living animal and plant cells and isthe primary tetrapyrrole precursor. It is available from a variety ofspecialty chemical sources, e.g., Sigma Chemical Co., St. Louis, Mo. andBiosynth International, Skokie, Ill. It is known that excised planttissues treated in the laboratory with small amounts of ALAwillsynthesize and accumulate Pchlide, which is the immediate precursor ofChlide a and of Chl a, and that ALA will induce the accumulation ofearlier tetrapyrrole intermediates of the Chl biosynthetic pathway, suchas coproporphyrin, Proto, and MP(E). Once the ALA has stimulated thesynthesis of the tetrapyrrole intermediates, they are normally convertedin the presence of sunlight into the various forms of Chl a, asdescribed in FIG. 1. However, this rate-limiting conversion does notoccur to any great extent in darkness; without sunlight, thetetrapyrrole intermediates accumulate in small amounts in theirrespective metabolic pools. Upon exposure to light, the conversion toChl a resumes and the pools are depleted. In 1974, Castelfranco, P. A.,Rich, P. M., and Beal, S. I., Plant Physiol. 53:615-618 noticed whilestudying the lag phase during greening of etiolated (dark grown) tissuethat excised cucumber cotyledons soaked in ALA for 16 hours in the darkunderwent visible tissue damage upon subsequent exposure to light, whichwas attributed to tetrapyrroles formed from exogenous ALA. Thisphenomenon was regarded as a nuisance to be avoided by illumination withred light of very low intensity or by illumination with intermittentlight. It was believed that the accumulation of tetrapyrroles due toexogenous ALA was a phenomenon attributable to the peculiarcircumstances of etiolation. Indeed, once the greening of etiolatedtissue is initiated, the biosynthesis of chlorophyll proceeds at anabnormally high rate not found in normal green tissue.

Copending application Ser. No. 895,529, the disclosure of whichapplication is expressly incorporated herein by reference, describesherbicidal compositions comprising one or more compounds selected fromthe group consisting of δ-aminolevulinic acid, inducers ofδ-aminolevulinic acid synthesis in plants, enhancers of δ-aminolevulinicacid conversion to photodynamic tetrapyrroles in plants, and inhibitorsof conversion of divinyl tetrapyrroles to monovinyl tetrapyrroles inplants; methods for inducing the accumulation of photodynamictetrapyrroles in living plants using said compositions, and methods ofcontrolling plants using said compositions. These compositions werediscovered to have a herbicidal effect on plants as the result of theaccumulation of tetrapyrroles in amounts greater than those normallyfound in the plants. This was surprising because mature green plantssynthesize chlorophyll only at a rate sufficient to keep up with leafexpansion and repair, and it had not been previously believed that thisrate would be sufficient to allow accumulation of amounts oftetrapyrroles large enough to result in photodynamic injury.

The accumulated tetrapyrroles photosensitize the formation of singletoxygen, which is a very strong oxidant. The singlet oxygen rapidlyoxidizes the lipoprotein components of the plant cellular membranes,thus setting in motion a highly destructive free-radical chain reaction,which can be summarized as follows (hv=photon of light; ¹Tet=tetrapyrrole in the singlet ground state; ³ Tet*=tetrapyrrole in thetriplet excited state; ³ O₂ =oxygen in the triplet ground state; ¹ O₂*=oxygen in the singlet excited state; UMLP=unsaturated membranelipoproteins):

(1) ¹ Tet+hv→³ Tet*

(2) ³ Tet*+³ O₂ →¹ Tet+¹ O₂ *

(3) ¹ O₂ *+(UMLP)→hydroperoxides

(4) hydroperoxides→free radicals

(5) free radicals+UMLP→more hydroperoxides

(6) repetition of steps (4) and (5) until most of the UMLP are oxidized.

While photosensitization by injected tetrapyrroles had been described inanimals and human tissues (see, e.g., Ellefson, R. D., Mayo Clinic Proc.57:454-458 (1982); Christensen, T., Sandquiet, T., Feren, K., Waksvik,H., and Moan, J., Br. J. Cancer 48:35-43 (1983); Hopf, F. R., andWhitten, D. G., in The Porphyrins, Vol. 2, Dolphin, D., ed. (AcademicPress, New York, 1978), pp. 161-195; Sandberg, S., Romslo, I., Hovding,G., and Bjorndal, T., Acta Dermatovener (Stockholm) Suppl. 100:75-80(1982); Latham, P. S., and Bloomer, J. R., Photochem. Photobiol.37:553-557(1983); Bickers, D. R., Dixit, R., and Mukhtar, H., Biochem.Biophys. Res. Comm. 108:1032-1039 (1982)), this phenomenon had not beendemonstrated in whole green plants nor adapted to control undesirablesusceptible plant species prior to the invention of Ser. No. 895,529.

OBJECTS OF THE INVENTION

It is therefore an object of the present invention to provide aphotodynamic composition which is capable of defoliating a plant withoutkilling the plant by causing the foliage to accumulate levels oftetrapyrroles which are higher than those normally found in the foliage.

It is a further object of the invention to provide a photodynamiccomposition for causing fruit drop in a plant, particularly in adeciduous fruit tree, without the need for mechanical harvesting.

It is yet another purpose of the invention to provide a photodynamiccomposition capable of causing defoliation and fruit drop in a plant.

It is another object of the invention to provide photodynamiccompositions which are capable of defoliating and/or defoliating andcausing fruit drop in a plant which are environmentally safe andefficient at low concentrations.

SUMMARY OF THE INVENTION

The invention is broadly directed to compositions for causingdefoliation and/or fruit drop in whole, living plants and methods fordefoliating and/or causing fruit drop in whole, living plants. Thus, inone embodiment, the invention is a plant defoliating compositioncomprising a plant defoliating effective amount of δ-aminolevulinic acidor δ-aminolevulinic acid in combination with one or more chlorophyllbiosynthesis modulators and a suitable carrier.

In another embodiment, the invention is a plant defoliating compositioncomprising a plant defoliating effective amount of δ-aminolevulinic acidin combination with one or more chlorophyll biosynthesis modulatorswhich are selected from the group consisting of inducers ofδ-aminolevulinic acid synthesis, enhancers of δ-aminolevulinic acidconversion to tetrapyrroles and inhibitors of conversion of divinyltetrapyrroles to monovinyl tetrapyrroles.

Another embodiment of the invention is a method for defoliating a plantcomprising the steps of contacting the plant with a defoliatingeffective amount of δ-aminolevulinic acid, one or more chlorophyllbiosynthesis modulators, or δ-aminolevulinic acid in combination withone or more chlorophyll biosynthesis modulators and allowing thecontacted plant to be exposed to light.

In another embodiment, the invention is a method for defoliating a plantcomprising the steps of contacting the plant with a defoliatingeffective amount of δ-aminolevulinic acid in combination with one ormore chlorophyll biosynthesis modulators and allowing the contactedplant to be exposed to light.

In still another embodiment, the invention is a method for defoliating aplant comprising the steps of contacting the plant with a defoliatingeffective amount of δ-aminolevulinic acid, one or more chlorophyllbiosynthesis modulators, or δ-aminolevulinic acid in combination withone or more chlorophyll biosynthesis modulators, exposing the contactedplant to a substantial absence of light at wavelengths of 300 to 700mMand then exposing the contacted plant to light.

A further embodiment of the invention is a composition for causingdefoliation and fruit drop in a deciduous fruit tree comprising anamount effective to cause defoliation and fruit drop in a deciduousfruit tree of δ-aminolevulinic acid or δ-aminolevulinic acid incombination with one or more chlorophyll biosynthesis modulators and asuitable carrier.

Another embodiment of the invention is a composition for causingdefoliation and fruit drop in a deciduous fruit tree comprising anamount effective to cause defoliation and fruit drop in a deciduousfruit tree of δ-aminolevulinic acid in combination with one or morechlorophyll biosynthesis modulators capable of causing the foliage ofthe tree to accumulate levels of tetrapyrroles which are selected fromthe group consisting of inducers of δ-aminolevulinic acid synthesis,enhancers of δ-aminolevulinic acid conversion to tetrapyrroles andinhibitors of the conversion of divinyl tetrapyrroles to monovinyltetrapyrroles.

In a specific embodiment, the invention is a composition for causingdefoliation and fruit drop in a deciduous fruit tree comprising anamount effective to cause defoliation and .fruit drop in a deciduousfruit tree of δ-aminolevulinic acid in combination with ethylnicotinate.

Still another embodiment of the invention is a method for causingdefoliation and fruit drop in a deciduous fruit tree comprising thesteps of contacting the tree with an amount effective to causedefoliation and fruit drop in the tree of δ-aminolevulinic acid, one ormore chlorophyll biosynthesis modulators, or δ-aminolevulinic acid incombination with one or more chlorophyll biosynthesis modulators andallowing the contacted tree to be exposed to light.

A further embodiment of the invention is directed to a method forcausing defoliation and fruit drop in a deciduous fruit tree comprisingthe steps of contacting a tree with an amount effective to causedefoliation and fruit drop in the tree of δ-aminolevulinic acid, one ormore chlorophyll biosynthesis modulators, or δ-aminolevulinic acid incombination with one or more chlorophyll biosynthesis modulators,exposing the contacted tree to a substantial absence of light atwavelengths of 300 to 700 mM and then exposing the contacted tree tolight.

DETAILED DESCRIPTION OF THE INVENTION

The invention will be understood more clearly and fully from thefollowing detailed description.

It has now been discovered that foliage of whole, living plants can beinduced by exposure to exogenous ALA to accumulate artificially highamounts of photodynamic tetrapyrrole intermediates in excess of levelsnormally found in foliage of living plants, and that such inducedartificially high levels are sufficiently photodynamic so thatsubsequent exposure of the foliage to light causes desiccation and deathof the foliage without killing the rest of the plant.

As a consequence of desiccation of the foliage of the plant, there isformation of an abscission layer between the branch and the leafpetioles (stems) which ultimately causes the leaves to fall from thebranch. Similarly, desiccation of the foliage of fruit bearing plants,e.g., deciduous fruit trees, causes formation of an abscission layerbetween the branch and the fruit pedicel (stem) or the pedicel and thefruit which ultimately causes the fruit Co drop from the branch.

The foliage of a plant can be induced to accumulate tetrapyrrolesbecause foliage is capable of synthesizing tetrapyrroles via thechlorophyll biosynthetic pathway. In contrast, "woody" parts of a plant,e.g., bark or stalk, do not actively synthesize tetrapyrroles and havesufficient carbohydrate reserves to recover from desiccation so that,notwithstanding desiccation of the foliage, the plant does not die.

As used herein, the term "plant" means a tree, shrub, seedling, or herb,which is a living organism and which typically does not exhibitvoluntary motion or possess sensory or nervous organs.

The term "woody" as used herein refers to plant tissue which does notactively synthesize tetrapyrroles and includes ligneous tissues, i.e.,tissue containing wood, wood fibers or wood-like fibers.

The term "herbaceous plant" refers to a plant having little or no woodytissue,

The term "deciduous tree" broadly refers to the opposite of evergreentree and includes trees whose leaves fall off seasonally or at a certainstage of development in the life cycle.

As used herein, the term "young leaf" refers to a leaf which is stillexpanding in size.

As used herein, the term "mature leaf" refers to a leaf which hasstopped expanding in size.

As used herein, the term "old leaf" refers to a mature leaf which isapproaching senescence.

As used herein, the term "desiccate" means broadly to dry and includesloss of cellular fluids, followed by degradation of chlorophyll (Chl)and other biomolecules such as proteins, lipoproteins, and nucleicacids, and disintegration of subcellular organelles such as vacuoles,nuclei, mitochondria, plastids, microsomes and microbodies.

As used herein, the term "defoliate" means broadly to remove green planttissue, in particular, foliage (leaves) and includes separation ofleaves at their junction to petioles, or separation of leaves andpetioles at their junction to branches, before or after complete leafdesiccation.

As used herein, the term "fruit drop" means broadly to remove fruit frombranches of plants, in particular deciduous fruit trees, and includesseparation of fruit at the junction to pedicels or separation of fruitand pedicels at their junction to branches before or after leafdesiccation.

As used herein, the term "fruit" means the mature ovary of a flowerwhich includes either parts of the flower or inflorescence which areintimately associated with the mature ovary. Fruit includes but is notlimited to apples, oranges, pears, peaches, cherries, tomatoes and thelike.

As used herein, the term "chlorophyll biosynthesis modulator" refers toa compound other than exogenous ALA (ALA from sources outside the plant)which causes the green tissue of a plant, e.g.., foliage, to accumulatelevels of tetrapyrroles which are higher than levels of tetrapyrrolesnormally found in untreated green tissue. Such modulators are selectedfrom the group consisting of inducers of ALA synthesis, enhancers of ALAconversion to tetrapyrroles and inhibitors of conversion of divinyltetrapyrroles to monovinyl tetrapyrroles. In accordance with theinvention, one or more modulators, or one or more modulators incombination with ALA can be used to effect defoliation or defoliationand fruit drop in plants.

Modulators of the invention include, e.g., o-phenanthroline,1,7-phenanthroline, 4,7-phenanthroline, and phenanthridine, availablefrom, e.g., Alpha Products, Danvets, Mass.; 2,2'-dipyridyl (2,2'-DP),2,3'-dipyridyl (2,3'-DP), 2,4'-dipyridyl (2,4'-DP), 1,7'-dipyridyl(1,7'-DP), 4,4'-dipyridyl (4,4'-DP), pyridine 2-aidehyde, pyridine2-aldoxime, 2,2'-dipyridylamine, 2,2'-dipyridyl disulfide,8-hydroxyquinoline, picolinic acid, nicotinic acid, 6-aminonicotinamide, ethyl nicotinate, 2-hydroxynicotinic acid, ethyl 2-methylnicotinate, N-methyl nicotinamide, N-benzyl-N-nicotinoyl nicotinamide,2-hydroxy-6-methylpyridine-3-carboxylic acid,4-hydroxy-7-trifluoromethyl-3-quinoline carboxylic acid,4-hydroxy-7-methyl-1,8-naphthyridine-3-carboxylic acid, diethyl3,4-pyridine dicarboxylate, and niflumic acid, available from, e.g.,Aldrich Chemical Co., Milwaukee, Wis.; and analogs thereof. Othermodulators are listed in Table XVI below. It should be noted thatnicotinic acid, an enzyme cofactor, occurs in all living cells andappreciable amounts are found in liver, yeast, milk, adrenal glands,white meat, alfalfa, legumes, whole cereals and corn. In addition, ethylnicotinate is a vitamin derivative.

By "inducer of ALA synthesis" is meant a compound which, when applied toplants, stimulates the green tissue of the plant to produce a higherthan normal amount of endogenous ALA ("native ALA," i.e., ALA normallyfound in a plant) which in turn causes accumulation of tetrapyrroles atlevels sufficiently photodynamic so that upon subsequent exposure of thetissue to light, the tissue desiccates. Thus, an inducer results in asignificant accumulation of a particular MV or DV tetrapyrrole whenapplied to a plant in the absence of exogenous ALA. Significantaccumulation of a particular tetrapyrrole is defined as an amount ofthat accumulated tetrapyrrole which approaches or exceeds the netdark-conversion rate into that tetrapyrrole, brought about by a 5 mMexogenous ALA treatment. Furthermore, the inducer, in combination withALA, results in the accumulation of higher levels of the particular MVor DV tetrapyrrole than when ALA or the inducer are applied to the plantseparately. Thus, the compositions of the invention can comprise one ormore inducers of ALA, or one or more inducers of ALA in combination withALA.

By "enhancer of ALA conversion to tetrapyrroles" or "enhancer" is meanta compound which when applied to plants enhances the capability of thegreen tissues of the treated plants to convert exogenous or endogenousALA to photodynamic tetrapyrroles. An enhancer does not result in asignificant accumulation of a particular MV or DV tetrapyrrole whenapplied to a plant in the absence of exogenous ALA but, when usedjointly with exogenous ALA, significantly enhances the dark conversionof exogenous ALA into the particular MV or DV tetrapyrroles over andbeyond that caused by exogenous ALA alone, i.e., by ALA used as acontrol. A significant accumulation of a particular tetrapyrrole in thiscontext is defined as the amount of that accumulated tetrapyrrole whichapproaches or exceeds the net dark-conversion rate into thattetrapyrrole brought about by a 5 mM exogenous ALA treatment. Enhancersof ALA conversion to tetrapyrroles fall into two groups: (1) enhancersof ALA conversion to MV Pchlide and (2) enhancers of ALA conversion toDV Pchlide and enhancers of ALA conversion to proto and MV- and DV-MPE.Thus, the compositions of the present invention can also comprise one ormore enhancers of ALA, or one or more enhancers of ALA in combinationwith ALA or inducers of ALA.

By "inhibitor of the conversion of divinyl tetrapyrroles to monovinyltetrapyrroles" is meant a compound which, when applied alone to plants,results in the inhibition of a particular MV tetrapyrrole in comparisonto untreated controls and/or when applied to a plant in combination withALA results in the inhibition of a particular MV tetrapyrrole incomparison to ALA-treated controls.

A modulator which functions as one of type of modulator (i.e., inducer,enhancer or inhibitor) in a specific plant or at a given concentrationmay function as a different type of modulator at a differentconcentration or in another plant, although a compound which is amodulator in one type of plant will be a modulator in most other typesof plants.

For example, 2,2'-dipyridyl can be an enhancer in cucumber atconcentrations less than 20 mM but can also be an inducer in cucumber atconcentrations of 20mM or greater. Further, in cucumber, a DDV/LDV plantspecies, 2,2'-dipyridyl and o-phenanthroline are inducers; pyridine2-aldoxime, pyridine 2-aidehyde, picolinic acid, 2,2'-dipyridyldisulfide, 2,2'-dipyridylamine, 4,4'-dipyridyl, phenanthridine,nicotinic acid, 2-hydroxynicotinic acid,2-hydroxy-6-methylpyridine-3-carboxylic acid, ethyl nicotinate,ethyl-2-methyl nicotinate and 4-hydroxy-7-trifluoro-8-quinolinecarboxylic acid are enhancers; and 2,3'-dipyridyl, 2-4'-dipyridyl,1,7-phenanthroline, 4,7-phenanthroline, diethyl 3,4-pyridinedicarboxylate and niflumic acid are inhibitors; in soybean, a DMV/LDVplant species, 2,4-dipyridyl, 2,2'-dypiridylamine, phenanthridine,picolinic acid, pyridine 2-aldoxime, 2,3-dipyridyl, 4,4'-dipyridyl,1,7-dipyridyl, pyridine 2-aidehyde, 2,2'-dipyridyl disulfide and8-hydroxyquinoline are enhancers; and 4,7-phenanthroline and1,7-phenanthroline are inhibitors; and in Johnsongrass, a DMV/LMV plantspecies, 2,2' -dipyridylamine, pyridine 2-aldoxime, pyridine 2-aidehyde,picolinic acid, 2,2'-dipyridyl, 2,4-dipyridyl, 1,7-phenanthroline,2,2'-dipyridylamine, 2,2'-dipyridyldisulfide, 2,3-dipyridyl and4,7-phenanthroline are enhancers; and 2,4-dipyridyl and 2,3-dipyridylare inhibitors. One skilled in the art will be able to determine,without undue experimentation, whether a compound is a modulator and, ifdesired, will be able to determine the type of modulator based on themethods disclosed herein.

Thus, the compositions of the present invention can also comprisecombinations of ALA and one or more chlorophyll biosynthesis modulatorsselected from the group consisting of inducers, enhancers, andinhibitors, e.g., ALA+one or more inducers, ALA+one or more enhancers,ALA+one or more inhibitors, ALA+one or more inducers+one or moreenhancers, ALA+one or more inducers+one or more inhibitors, ALA+one ormore enhancers+one or more inhibitors, ALA+one or more inducers+one ormore enhancers+one or more inhibitors, etc.

A consideration of one or more of the following factors will enable oneskilled in the art to effect the desired defoliation and/or fruit dropfor a given plant species: the species of the plant (monocot, dicot,annual, perennial, woody, non-woody); the age of the plant; the varioustissues types present on the plant (cotyledons; stems, leaves, leafpetioles, growing points, fruit pedicels, bark, etc.); and the point oftime in the growing season. For example, (a) spraying a plant with woodybranches will result in the desiccation of the green leaves but not thewoody branches because the woody branches are protected by suberizedbark which does not respond to treatment by accumulating tetrapyrroles;(b) spraying a young plant with tender, succulent stems containingchlorophyll will desiccate both the leaves and the stems, whiletreatment of plants with branches protected by suberized bark willresult in desiccation of the leaves only; (c) spraying stems containinggreen leaves and unprotected growing points (e.g., leaf and flower buds)will desiccate both the leaves and the growing points, while sprayingstems with leaves and growing points protected by suberized scales willonly desiccate the leaves leaving the protected growing pointsunaffected; (d) spraying plants with young and old leaves may result inthe desiccation of a larger proportion of the old or young leaves,depending on the nature of the modulator (i.e., inducer, enhancer orinhibitor) used with ALA; (e) spraying an annual plant with fewcarbohydrate reserves will result in desiccation followed by a slowerrebound than a perennial plant with more carbohydrate reserves; and (f)spraying a woody plant, with carbohydrate reserves stored in the woodystems and roots, at the end of the growing season will result indesiccation of the leaves without resprouting of new leaves, whilespraying the same plant early in the growing season will result indesiccation of the treated leaves, but with regeneration of new leavesfrom the carbohydrate reserves stored in the stems and roots, givenproper temperature and daylength conditions.

The compositions of the present invention can contain one or more of thefollowing: suitable carrier(s) (e.g., colloidal magnesium aluminumsilicate, pumice, talc, or combinations thereof); solvent(s) (e.g.,water, 0.45 acetone: 0.45 ethanol:0.1 Tween 80:9 water (v/v/v/v), 0.45acetone:0.45 methanol:0.1 Tween 80:9 water (v/v/v/v), 0.1-1% Tween 80 inwater (v/v), 0.9 polyethylene glycol (PEG):0.1 Tween 80:9 water (v/v/v),0.1-0.7 PEG:0.2--0.8 methanol:0.1 Tween 80:9 water (v/v/v/v), 0.9methanol:0.1 Tween 80:9 water (v/v/v), 0.45 acetone:0.45 ethanol:0.2Tween 80:0.9 ethylene glycol:18 water (v/v/v/v/v), or one or more of thefollowing: benzene, toluene, xylene, kerosene, 2-methoxyethanol,propylene glycol, diethylene glycol, diethylene glycol diethyl ether,formamide, methylformamide, cyclohexanone, isophorone); buffer(s) (e.g.,citric acid); wetting agent(s) (e.g., sodium N-methyl-N--oleoyltaurate,an alkylphenoxy polyoxyethylene ethanol, sodium olefin sulfonate, sodiumisopropylnaphthalene sulfonate, polyoxyethylated vegetable oil);dispersing agent(s) (e.g., sodium lignin sulfonate, the sodium salt of anaphthalene sulfonic acid-formaldehyde condensate, hydroxyethylcellulose); defoaming agent(s) (e.g., silicone); emetic(s) (e.g., sodiumtripolyphosphate, tetra potassium pyrophosphate, arecotine, apomorphine,copper sulfate); stench(es) (e.g., pyridine); penetrant(s);surfactant(s); emulsifier(s); and adjuvant(s) (e.g., phytoblend oils).Of course, any such additional component must be compatible with theactive ingredients of the present invention and with the otheringredients in the mixture.

The compositions can be formulated in any manner conventionally used forplant preparations, e.g., as a solution, suspension, emulsion, flowableconcentrate, emulsifiable concentrate, gel, paste, foam, cream, aerosol,wettable powder, dust, dispersible granules, and the like, according toprocedures known to those skilled in the art. Advantageously, thecomposition is a solution, suspension, emulsion, aerosol, flowable oremulsifiable concentrate, or wettable powder. Of course, the formulationmust be such that the active ingredient(s) penetrate(s) the plant tissueand translocates to the sites of tetrapyrrole synthesis. When thecompositions are made in solution they can conveniently compriseconcentrations of from about 1 to about 40 mMALA, advantageously, 15 mMto 40 mM, and from about 5 to about 30 mM inducer, enhancer, orinhibitor, advantageously 15 to 30 mM.

The compositions of the present invention can be applied topically,e.g., as a dust, soak, dip, spray, mist, or fog, in an amount sufficientto induce the accumulation of photodynamic tetrapyrroles. Alternatively,the compositions can be applied to the soil for uptake by plant rootsand translocation to the vegetative part of the plant. The amount ofcomposition to be applied will vary, depending on the particular activeingredient(s) selected, but in general will be an amount sufficient tosupply from about 10 g to about 15 kg ALA per acre and/or from about 10g to about 10 kg of an inducer, enhancer, or inhibitor per acre. Meansof determining optimum application rates are within the purview of thoseskilled in the art.

Once the tissues of the plant have been induced to begin accumulatingartificially high amounts of tetrapyrroles by exposure to thecompositions of the present invention, the plant may be shielded fromexposure to light to allow maximum tetrapyrrole accumulation. Such darkincubation is not required for activity but tends to optimize efficiencyof the compositions. The plants can be shielded in any convenientmanner, as by wrapping them in dark paper, cloth, or foil, or by placingthem in a dark room or container. Under field conditions, the idealmethod to provide a period of dark incubation is to apply thecomposition at dusk or during the night, at a time chosen to allow theplants to rest in the dark for at least one hour. It is to be understoodthat in order to facilitate tetrapyrrole accumulation, the dark need notbe total absence of light, but rather substantial absence of light atwavelengths of from 300 to 700 nm. Advantageously, the plants areallowed to rest in the dark for from about 1 to about 20 hours. One to 8hours is particularly advantageous.

Thereafter the plants are exposed to about 200 ft. candles or more oflight at wavelengths of about 300 to about 700 nm. The light can besupplied by any convenient source, e.g., an incandescent lamp, metalhalide lamp, sunlamp, or a cool white or skylight fluorescent bulb. Inthe field, of course, the preferred source of light is sunlight. Theplants are exposed to light for a period of time sufficient to oxidizemost of the unsaturated membrane lipoproteins; a period of from about 1to about 14 days is preferred.

A further understanding of this invention can be had from the followingillustrative examples. Desiccating activity is indicated by tissuenecrosis and leaf abscission and/or fruit drop. As used hereinabove andbelow unless expressly stated to the contrary, all temperatures andtemperature ranges refer to the centigrade system and the terms ambientand room temperature refer to about 20°-25° C. The term percent or (%)refers to weight percent and the terms mole and moles refer to grammoles. "Level of significance" refers to the probability that for apopulation for which the correlation coefficient (r) is equal to zero, asample of size n can be taken, for which the correlation coefficientequals or exceeds the calculated value of r reported for the givensample. The abbreviation "n.s." stands for "not significant".

SECTION I PROTOCOL FOR DETERMINING PHOTODYNAMIC DEBICCATING COMPOSITIONS

The following examples describe model systems whereby persons skilled inthe art can readily determine photodynamic compounds and compositionsuseful in the present invention.

EXAMPLE 1 photodynamic Effects of ALA

Cucumber (Cucumis sativus L. cv Beit Alpha MR) seedlings were germinatedin the greenhouse in vermiculite in glass containers, 9 cm deep and 9 cmin diameter. The seedlings were watered periodically with Hoaglandsolution. The photoperiod was maintained at 14 hours of light per daywith 50 ft. candles of incandescent light.

Six-day old green seedlings were thinned to 10 plants per container andALA (Sigma Chemical Co., St. Louis, Mo.) was applied as a fine spray.The ALA was dissolved at concentrations ranging from 0 to 20 mM in asolvent mixture made up of 0.45 acetone:0.45 ethanol:0.1 Tween 80:9water (v/v/v/v), adjusted to pH 3.5 with dilute HCl. Each 9 cm-diameterglass container (approximately 63.6 cm² leaf surface area) was sprayedwith 0.25 ml of ALA (treated) or 0.25 ml of solvent (control), which isequivalent to a spray rate of about 40 gallons/acre and a fieldapplication rate of ALA of about 0 to 524 g/acre. The solutions weredelivered as a very fine and uniform spray with a modified Pierce"Quixspray" aerosol spray kit (Pierce Chemical Co., Rockford, Ill.), asfollows: 0.25 ml of solution was placed in a sawed-off 10 ml conicalcentrifuge tube, which was placed inside the Quixspray spray jar. Thedelivery of a very fine mist was achieved by pumping the solutionthrough a fine bore polypropylene tubing (0.3 mm inside diameter, or 0.5mm inside diameter for more viscous solutions). One end of the fine-boretubing was inserted into the Quixspray intake hose, while the other endwas dipped into the solution in the conical centrifuge tube. In thismanner it took 10-20 sec to deliver 0.25 ml spray, and this in turnprovided ample time for thoroughly spraying the seedlings to leafsaturation. Each treatment was performed in duplicate. Average dropletsize diameter was approximately 25 μm for the 0.3 mm tubing and about 50μm for the 0.5 mm tubing.

After spraying, the plants were wrapped in aluminum foil and were placedinside a cardboard box which was wrapped in two layers of black plastic.The dark-boxes were then incubated overnight (17 hours) at 28° C., inorder to allow the biosynthesis and accumulation of tetrapyrroles totake place.

The next morning, the treated plants were sampled for their tetrapyrrolecontent. The plants were taken in the black boxes to a dark roomequipped with a green safelight which permits the manipulation of thetreated tissues without affecting in any way their tetrapyrrole content.One of each two cotyledons of every two replicates was excised. Two- tothree-gram batches were then homogenized in a Sorval Omnimixer (DuPontInstruments, Newtown, Conn.) in acetone:0.1N NH₄ OH (9:1 v/v) at a rateof 18 ml of solvent per 3 g of tissue. The resulting 80% acetone extractcontaining various tetrapyrroles was cleared from lipoproteins and celldebris by centrifugation at 39,000 × g for 10 min at 0° C. Chlorophyll,a fully esterified tetrapyrrole, was removed from the aqueous acetonesolution by extraction with hexane according to the method of Rebeiz, C.A., Mattheis, J. R., Smith, B. B., Rebeiz, C. C., and Dayton, D. F.Arch. Biochem. Biophys. 166:446-465 (1975). The more polar mono- anddicarboxylic tetrapyrroles such as Proto, MP(E), and Pchlide remained inthe hexaneextracted aqueous acetone fraction. The chemical structure ofthese tetrapyrroles has been discussed at length in Rebeiz, C. A. andLascelles, J., in Photosynthesis: Energy Conversion by Plants andBacteria, Vol. 1, Govindjee, ed. (Academic Press, New York, 1982), pp.699-780; and Rebeiz, C. A., Wu, S. M., Kuhadja, M., Danjell, H., andPerkins, E. J. Mol. Cellular BioChem. 57:97-125 (1983). The amount ofProto, MP(E), and Pchlide was determined spectrofluorometrically onaliquots of the hexane-extracted acetone fraction according to themethod of Rebeiz, C. A., Mattheis, J. R., Smith, B. B., Rebeiz, C. C.,and Dayton, D. F., Arch. Biochem. Biophys. 171:549-567 (1975). A smallaliquot of the hexane extract containing the Chl a and b was dried underN₂ gas and the residue was redissolved in 80% acetone. The amount of Chla and b in this acetone solution was then determinedspectrofluorometrically according to the method of Bazzaz, M. B., andRebeiz, C. A., Photochem. Photobiol. 30:709-721 (1979).

Fluorescence spectra were recorded on a fully corrected photon countingspectrofluorometer Model SLM 8000 DS (SLM-Aminco, Urbana, Ill.) equippedwith two red-sensitive, extended S₂₀ photomultipliers (EMI 9658), andinterfaced with a microcomputer system Model 9825 S (Hewlett-Packard,Sunnyvale, Calif.). Tetrapyrrole solutions were monitored at roomtemperature on 0.3 ml samples, in cylindrical microcells, 3 mm indiameter. Conversion of the digital spectral data into concentrationswas performed automatically by the microcomputer, following therecording of the pertinent spectra, according to the method of Rebeiz,C. A., Daniell, H., and Mattheis, J. R., Biotech. Bioeng. Symp. No.12:413-439 (1982). The emission and excitation spectra were recorded atexcitation and emission bandwidths of 2 mm.

Monovinyl tetrapyrroles were distinguished from divinyl tetrapyrroles bytheir well-established spectrofluorometric properties in ether at 77 °K. (see Rebeiz and Lascelles, supra; Rebeiz, Wu, Kuhadja, Daniell andPerkins, supra; Belanger, F. C., and Rebeiz, C. A., J. Biol. Chem.257:1360-1371 (1982); and Belanger, F. C., Duggan, J. X., and Rebeiz, C.A., J. Biol. Chem. 257:4849-4858 (1982)). The low temperaturefluorescence emission and excitation spectra were recorded incylindrical sample tubes as described in Cohen, C. E., and Rebeiz, C.A., Plant Physiol. 61:824-829 (1978).

Absorption spectra were recorded with an Aminco dual wavelengthspectrophotometer model DW-2 (SLM-Aminco, Urbana, Ill.) operated in thesplit-beam mode, at a slit width of 2 nm.

The acetone-insoluble residue which was left behind after centrifugationof the tissue homogenate was suspended in distilled water with an allglass tissue grinder. Total proteins were determined on a small aliquotof the suspension, after delipidation, according to the method ofRebeiz, C. A., Castelfranco, P. A., and Engelbrecht, A. H., PlantPhysiol. 40:281-286 (1965).

After 17 hours of dark- incubation the treated plants accumulated 382.82and 2.36 nmoles of Pchlide and MP(E), respectively, per 100 mg protein,above and beyond the controls. The seedlings with half of theircotyledons still intact were then used for assessing photodynamic damageby light. The seedlings were exposed to daylight in the greenhouse (400to 5000 ft. candles at noon, depending on cloud cover) and their growthwas evaluated over a period of 10 days. In order to secure a permanentrecord of the growth behavior of the treated plants, the latter werephotographed daily (Kodacolor, 400 ASA, Eastman Kodak Co., Rochester,N.Y.) with a Pentax Super Program camera (Helix, Champaign, Ill.)equipped with an SMC Pentax-A 1:1.4 50 mm lens and a digital back thatimprinted on each photograph the date or time of day at which thephotograph was taken. Per cent photodynamic damage was assessed as thepercent death of the sprayed tissue, in response to exposure tosunlight. For example, if 10 out of 10 sprayed leaves or cotyledons diedas a consequence of exposure to daylight, the photodynamic damage wasconsidered to be 100%. If only five out of the ten sprayed leaves orcotyledons had died, the photodynamic damage was considered to be only50%, etc.

The extent of photodynamic damage was related to the amount ofaccumulated tetrapyrroles by conventional correlation analysis. Theamounts of tetrapyrrole that accumulated were expressed in nmoles per100 mg of tissue protein.

The results of these experiments are shown in Table I.

                                      TABLE I                                     __________________________________________________________________________    Various Concentrations of ALA                                                                change.sup.2 after 17 h of dark-incubation                     Experi-                                                                           Treatment  nmol/100 mg protein Photodynamic                               ment.sup.1                                                                        mM ALA                                                                              g/acre                                                                             Pchlide                                                                           MP(E)                                                                             Proto                                                                             Tetrapyrroles.sup.3                                                                   damage (%)                                 __________________________________________________________________________    A   0(Control)                                                                           0   0.00                                                                              0.00                                                                              0.00                                                                              0.00     0                                              5    131  31.77                                                                             -4.44                                                                             -17.33                                                                            10.00   22                                             10    262  132.76                                                                            -0.50                                                                             -13.07                                                                            119.19  45                                             15    393  271.29                                                                            1.23                                                                              -17.33                                                                            255.19  95                                             20    524  210.60                                                                            -0.75                                                                             -17.33                                                                            192.52  85                                         Correlation coefficient                                                                      0.988       0.978                                              Level of significance                                                                        0.1%        0.1%                                               B   0(Control)                                                                           0   0.00                                                                              0.00                                                                              0.00                                                                              0.00     0                                              1     26  24.95                                                                             -3.08                                                                             -2.81                                                                             61.12    2                                              5    131  140.82                                                                            -1.69                                                                             -12.41                                                                            123.72  63                                             10    262  147.82                                                                            3.83                                                                              19.58                                                                             168.23  92                                             15    393  191.22                                                                            0.78                                                                              -12.41                                                                            175.03  95                                         Correlation coefficient                                                                      0.998       0.975                                              Level of significance                                                                        0.1%        0.1%                                               __________________________________________________________________________     .sup.1 In experiment A, the light intensity at noon during the first day      of exposure to daylight was about 400 ft. candles. In experiment B, it wa     about 5000 ft. candles.                                                       .sup.2 The change in tetrapyrrole concentration is the difference between     the tetrapyrrole content of the ALAtreated plants and that of the control     plants which were sprayed with the solvent only, i.e. without added ALA,      after 17 h of dark incubation and just prior to exposing the plants to        daylight. The control plants contained the following amounts of               tetrapyrrole after 17 h of dark incubation, and prior to exposure to          daylight: A: 99.6, 7.66, 17.33 and B: 22.69, 6.96, and 12.41 nmo le           Pchlide, MP(E) and Proto respectively per 100 mg protein.                     .sup.3 = Pchlide + MP(E) + Proto                                         

The symptoms of photodynamic damage assumed two forms: bleaching of thegreen leafy tissue, which spread gradually; and severe bleaching of thehypocotyl. In both cases, this was accompanied by a severe loss ofturgidity of the affected tissues. The photodynamic damage was effectedon the cell membranes which became leaky and this in turn resulted in arapid and severe dehydration of the tissues. For example, at ALAconcentrations of 10-20 mM (262-524 g/acre) a large number of seedlingshad undergone irreversible damage after four to five hours of exposureto daylight. The cause of death was usually due to severe dehydration,bleaching, and collapse of the leafy and/or hypocotyl tissues. On theother hand, treated samples kept for the same period of time in darknesswere unaffected.

EXAMPLE 2 Photodynamic Response of Various Plant Species to ALA+2,2'-DFTreatment

The procedure of Example I was performed on the following representativemonocots and dicots:

Cucumber (Cucumis Sativus L. cv Beit Alpha MR)

Lambsquarter (Chenopodium album)

Mustard (Brassica kaber/juncea)

Red root pigweed (Amaranthus retroflexus)

common purslane (Portulaca oleracea)

Tomato (Lycopersicon esculentum cv Jet Star)

Cotton (Gossypium herbacium cv Coker-315)

Red kidney bean (Phaseolus vulgaris L. cv. California Dark Red)

Soybean (Glycine max cv Williams)

Perennial bluegrass (Poa pratensis cv Aspen)

Barley (Hordeum vulgare, var. Beacon Spring)

Sweet corn (Zea mays L. cv Gold Cup)

Crabgrass (Digitaria sanguinalis L. and Digitaria ischaemum)

Giant foxtail (Setaria faberii)

Oat (Arena satira cv Centennial)

Wheat (Triticum sativum cv Auburn)

The greenhouse-grown seedlings were treated with 0.25 ml of 5 mM (131g/acre) ALA+15 mM (402 g/acre) 2,2'-DP, pH 3.5. Controls were treatedwith solvent only. All plants were then incubated in the dark for 17hours. The next morning the seedlings were sampled in the dark fortetrapyrrole content using the procedure of Example I for dicots and thefollowing procedure for monocots: the seedlings of one of the tworeplicates were excised into an upper half and a lower half. The twobatches of excised tissue were then homogenized separately in a SorvalOmnimixer in acetone:0.1N NH₄ OH (9:1 v/v) at a rate of 18 ml of solventper 3 g of tissue. The other replicate was used to assess thephotodynamic effect of light on the seedlings. For some dicots, thestems as well as the leaves were analyzed for tetrapyrroles. The resultsare given in Table II:

                                      TABLE II                                    __________________________________________________________________________    Photodynamic Response of Various Plant Species to ALA + 2,2'-DP Spray                     Age at                                                                             Type of         nmol/100 mg protein Photodynamic                         spraying                                                                           herbicidal                                                                          PChlide   MP(E)     Proto     damage (%)               Plant       (days)                                                                             response.sup.1                                                                      Control                                                                            Treated                                                                            Control                                                                            Treated                                                                            Control                                                                            Treated                                                                            Control                                                                            Treated             __________________________________________________________________________    Cucumber cotyledons                                                                        6   I     84.79                                                                              434.12                                                                             8.51 68.85                                                                              3.64 19.06                                                                              0    85                  Cucumber stems                                                                             6   I     10.87                                                                              71.77                                                                              5.32 14.47                                                                              12.67                                                                              39.37                                                                              0    85                  Lambsquarter                                                                               7   I     23.52                                                                              72.58                                                                              3.83 33.94                                                                              17.59                                                                              13.87                                                                              0    100                 Mustard leaves                                                                            12   I     29.84                                                                              200.82                                                                             12.01                                                                              36.11                                                                              29.08                                                                              23.52                                                                              0    90                  Mustard stems                                                                             12   I     15.26                                                                              49.60                                                                              2.95 13.13                                                                              0.00 38.35                         Red root pigweed                                                                          11   I     29.47                                                                              59.08                                                                              1.64 20.59                                                                              0.00 2.90 0    95                  Common purslane                                                                           21   I     8.37 33.30                                                                              1.54 11.79                                                                              1.88 5.71 0    80                  Tomato cotyledons                                                                         13   I     27.19                                                                              114.86                                                                             0.69 34.40                                                                              0.31 0.31 0    90                  Tomato stems                                                                              13   I     3.69 14.26                                                                              0.82 2.53 0.00 0.00 0    90                  Cotton cotyledons                                                                         14   II    18.06                                                                              36.53                                                                              3.95 9.22 0.00 0.00 0    63                  Cotton stems                                                                              14   II    3.70 4.18 1.19 1.13 0.00 0.00 0    0                   Kidney bean leaves                                                                         9   II    117.03                                                                             438.79                                                                             3.11 430.12                                                                             4.88 21.42                                                                              0    100                 Kidney bean stems                                                                          9   II    36.78                                                                              82.26                                                                              3.89 75.90                                                                              3.49 14.2 0     0                  Soybean leaves                                                                             9   II    25.31                                                                              98.88                                                                              3.61 105.84                                                                             4.24 10.87                                                                              0    78                  Soybean stems                                                                              9   II    6.06 6.17 0.37 0.45 0.00 0.45 0     0                  Perennial bluegrass                                                                       18   II    9.87 39.46                                                                              0.39 43.52                                                                              0.54 51.46                                                                              0    30-40               Barley       6   III   12.69                                                                              58.64                                                                              0.8  3.39 0.39 1.11 0    S.N..sup.2          Corn         9   III   79.09                                                                              85.44                                                                              4.90 15.47                                                                              12.39                                                                              0.00 0    S.N.                Crabgrass   25   III   44.43                                                                              114.32                                                                             3.13 27.63                                                                              0.00 0.00 0    S.N.                Giant foxtail                                                                              6   III   7.87 78.75                                                                              0.44 11.91                                                                              0.00 13.92                                                                              0    S.N.                Oat upper half                                                                             7   III   29.19                                                                              171.96                                                                             13.02                                                                              23.04                                                                              0.00 0.00 0    S.N.                Oat lower half                                                                             7   III   92.53                                                                              121.86                                                                             9.37 3.84 0.00 0.00 0    0.0                 Wheat upper half                                                                           7   III   29.58                                                                              101.25                                                                             8.34 5.22 9.96 0.60 0    S.N.                Wheat lower half                                                                           7   III   31.87                                                                              47.23                                                                              2.10 0.99 0.00 0.00 0    0.0                 __________________________________________________________________________     .sup.1 These types of photodynamic response are discussed below.              .sup.2 S.N. = small necrotic areas.                                      

An examination of results of this survey revealed that plants reacted inthree different ways to the ALA+2,2'-DP spray. One group of dicots,which is exemplified by cucumber, exhibited what is referred to as TypeI herbicidal response in Table II. This group of plants reacted to theALA+2,2'-DP spray exactly as did cucumber. Leafy tissues, stems andgrowing points accumulated significant amounts of tetrapyrroles and weresubject to severe photodynamic damage (Table II). Usually, the seedlingsdied very rapidly, and the rapidity of the response depended on thelight intensity in the greenhouse. For example, at the low sprayconcentrations used in this work (131 g/acre ALA+402 g/acre 2, 2'-DP),only 4 to 5 hours of exposure to daylight was sufficient to cause thedeath of the plants on clear, bright days (4000 to 6000 ft. candles atnoon). On the other hand, 2 to 3 days of insolation were required onvery cloudy days (400 ft. candles at noon) in order to achieve the sameresults. Some of the plant species that exhibited this type ofphotodynamic herbicidal response such as lambsquarter, mustard, redrootpigweed and common purselane are considered to serious weeds. While13-day old tomato plants, with fully expanded cotyledons and with smalldeveloping primary leaves exhibited a Type I response (Table II),younger 8- to 10-day old tomato seedlings were much less affected by thespray (approximately 40% photodynamic damage).

Other dicots such as cotton, kidney bean and soybean exhibited adifferent response to the ALA+2,2-DP treatment. This response isreferred to as Type II in Table II. Plants belonging to this groupaccumulate significant amounts of tetrapyrroles in the leafy tissues,but not in the stems as in cotton and soybean. Other species such askidney bean also accumulated some tetrapyrroles in the stems. Leavesthat accumulate tetrapyrroles exhibit very severe photodynamic damageand die within a few hours. However, the cotyledons, stems, and growingpoints remain unaffected. Such plants usually recovered from theoriginal photodynamic damage by producing new leaves and may require asecond application. In this group the Type II response also depended onthe age of the seedlings. For example, 6-day old soybean in which theprimary leaves were still enclosed within the cotyledons were completelyunaffected by the ALA+2,2 '-DP treatment. On the other hand, 9-day oldsoybean plants, with expanded primary leaves, exhibited a typical TypeII photodynamic herbicidal response. The only monitored monocot thatexhibited this type of response was perennial blue grass in which about30-40% of the sprayed leaves died; the plants subsequently recovered anddeveloped new leaves.

The third type of photodynamic herbicidal response elicited by theALA+2,2'-DP treatment is referred to as a type III response. Based onavailable data, monocots exhibited this type of response. Although theALA+2,2'-DP treatment induced the accumulation of significant amounts oftetrapyrrole by the plants, the photodynamic damage was eitherimperceptible as in wheat, oat, and corn, or when noticeable as inbarley, was confined to the upper half of a small proportion of thesprayed plants. In that case the photodynamic damage consisted of smallnecrotic regions. The seedlings continued to grow vigorously anddeveloped into healthy plants.

The photodynamic formulations described in this example exhibited anexcellent measure of species, age and organ-dependent selectivity. Whiledicotyledenous weeds such as lambsquarter, mustard, red root pigweed andcommon purselane were highly susceptible to the tetrapyrrole-inducedphotodynamic damage, monocots such as corn, wheat, oats, and barley werenot adversely affected by the spray. Other dicots were either unaffectedby the spray at an early stage of development as in soybean, orrecovered fully from a rapid destruction of the primary leaves byproducing new and healthy leaves, as was observed for kidney bean,soybean and cotton. Furthermore some tissues which accumulatedtetrapyrroles such as bean stems did not exhibit any photodynamicdamage. The biochemical basis of this organ, age and species-dependentphotodynamic herbicidal selectivity appears to be dependent among otherthings on the rates of tetrapyrrole turnover and on a differentialenhancement of the MV and DV tetrapyrrole biosynthetic pathways in anygiven plant species.

SECTION II EXAMPLES OF DEFOLIATION AND FRUIT DROP EXAMPLE 3 DefoliationOf "Red Delicious", "Golden Delicious", "Winesap" and "Prima" AppleCultivars Under Greenhouse Conditions

Scion wood of "Red Delicious", "Golden Delicious", "Winesap" and "Prima"apple cultivars was collected from the University of Illinois PomologyResearch Farm. These were "whip and tongue" grafted onto seedlingrootstock (purchased from Pacific Coast Nursery, Portland, Oreg.).Following grafting, seedlings were placed in plastic bags containingmoist vermiculite, and kept in a cold room for two weeks at 1° C. toinduce healing. The seedlings were then held in a cold chamber at 10° C.until needed.

Seedlings were later planted in 15 cm plastic pots containing a 1:1:1:1(v/v/v/v) mixture of soil, vermiculite, peat, and sand media. These weregrown in a greenhouse at 28° C. under a 14 h light--10 h darkphotoperiod. Light intensity was supplied by three, 1000 W metal halidelamps. Light intensity was 24 W/m².

Following five days of growth at 23° C., the apple seedlings weretreated as follows: (a) solvent only (control); (b) 20 mM ALA (BiosynthInt'l, Skokie, Ill.); (c) 30 m Methyl nicotinate (EN) (Aldrich Chem.Co., Milwaukee, Wis.); and (d) 20 mM of ALA plus 30 mM of EN. Thesolvent consisted of polyethylene glycol (Sigma Chem. Co., St. Louis,Mo.):methyl alcohol:Tween 80:water at 7:2:1:90 (v/v/v/v) (Rebeiz, C. A.,Montazer-Zouhoor, A., Mayasich, J. M., Tripathy, B. C., Wu, S., andRebeiz, C. C., CRC Crit. Rev. in Plant Sci., 6:385-435 (1988)). pH wasadjusted to 3.5 to facilitate the penetration of ALA into the leaves.Solutions were delivered as a fine and uniform spray with a spray gunnozzle. The solutions were placed in a Binks Wren air brush, and thedelivery of a fine mist (average diameter droplet size of 125 microns)was achieved by pumping the solution through metal tubing usingcompressed CO₂. One end of the tube was inserted into the gun intakehose, while the other end was dipped into the solution. Leaves weresprayed to a drip.

Treatment of the four apple cultivars was replicated three times in arandomized split plot design (four trees per replicate of eachcultivar). After spraying, trees were transferred to a dark-growthchamber at 28° C. and kept from 6:00 p.m. to 9:00 a.m. the followingmorning in order to allow for the dark conversion of ALA totetrapyrroles.

After 15 hours in darkness, plants were moved to the greenhouse forevaluation of growth and photodynamic damage over a period of 30 days.Plants were periodically photographed to record response to treatment. APentax Super program camera, equipped with an SMC Pentax-A 1:1.4, 50 mmlens, and a digital back recording the date and time of each shot on theprints obtained, was used. A 400 ASA Kodacolor film was used. Firstphotographic records were taken before treatment and then at 1, 3, 8,10, 17 and 30 days after exposure to light. Photodynamic damageconsisted of the sum of tissue necrosis and leaf abscission.

A gram leaf sample (collected from both the top and the bottom of eachof the seedlings) was homogenized in a Polytron homogenizer for oneminute in 7 ml of acetone and 0.1N ammonium hydroxide (9:1, v/v).Acetone serves to extract the tetrapyrroles while ammonium hydroxidemaintains the medium basic, thereby preventing loss of the Mg-atom frommetalated tetrapyrroles. During homogenization, samples were handledunder a green safelight which does not affect the tetrapyrrole contentof the tissues.

After homogenization, extracts were centrifuged at 18,000 rpm for 12min. at 1° C. to separate lipoproteins and cell debris from thesupernatant containing tetrapyrroles. Chlorophyll and other fullyesterified tetrapyrroles were removed by first extracting thesupernatant with hexane followed by an equal volume of hexane and thenwith a 1/3 volume of hexane (Rebeiz, C. A., Mattheis, J. R., Smith, B.B., Rebeiz, C. C., and Dayton, D. F. Arch. Biochem. Biophys. 171:549-567(1975)). Protoporphyrin IX (proto), Mg-protoporphyrin monoester (MPE),and protochlorophyllide (Pchlide) remained in the hexane-extractedacetone fraction (HEAF). The amount of tetrapyrroles in the HEAF wasdetermined by spectrofluorometry.

Fluorescence emission and excitation spectra of the HEAF were recordedat room temperature (23.5° C.) and at 77° K. (frozen in liquid nitrogen)using a fully corrected photon counting spectrofluorometer (model SLM8000 DS) equipped with two red-sensitive, extended S 20 photomultipliers(EMI 9658), and interfaced with an IBM Model 30 PC. Room temperaturedeterminations were performed on 0.3 ml aliquots in a cylindricalmicro-cell of 3 mm in diameter. Analytical techniques used for precisequantitative determination of the various tetrapyrroles are described byRebeiz, C. A., Mattheis, J. R., Smith, B. B., Rebeiz, C. C., and Dayton,D. F., Arch. Biochem. Biophys. 171:549-567 (1975); Smith, B. B., andRebeiz, C. A., Photochem. Photobiol. 26:527-532 (1977); Bazzaz, M. B.,and Rebeiz, C. A., Photochem. Photobiol. 30:709 (1979); and Rebeiz, C.A., Danjell, H., and Mattheis, J. R., in 4th Symp. Biotechnol. EnergyProd. Conserv. (Scot, C. D., Ed.) John Wiley & Sons, New York, 413-439(1982). Recording of spectra and conversion of digital spectral datainto concentrations were automatically performed by the microcomputer(Rebeiz, C. A., Danjell, H., and Mattheis, J. R., supra). For roomtemperature spectra, both the emission and the excitation band widthwere set at 4 nm. The photon count was integrated for 0.5 seconds ateach 1 nm increment. Ratios of MV MPE to DV MPE and of MV pchlide to DVPchlide were calculated from spectra recorded in ether at 77° K.(Tripathy, B. C., and Rebeiz, C. A., Anal. Biochem. 149:43-61 (1985)).The MV and DV tetrapyrrole content in a sample was determined from MVand DV tetrapyrrole ratios obtained at 77° K. and from total pigmentcontent determined at room temperature.

Total protein was determined using the bicinchoninic acid (BCA) methodSmith, P. K., Krohn, R. I., Hermanson, G. T., Mallia, A. K., Gardner, F.H., Provenzano, M. D., Fujimoto, E. K. Goeke, N. M., Olson, P. J. andKlenk, D. C., Anal. Blochem. 150:76-85 (1985), after resuspending thepellet obtained after centrifugation in distilled water. Absorbance wasmonitored on a Sequoia-Turner Model 340 spectrophotometer. Statisticalanalysis was conducted on an IBM P.C. Model 80, using a software packageavailable from SAS Industries, Inc. Data were analyzed as a randomizedsplit block design using the linear model. Testing for significantdifferences among cultivars and/or treatments was evaluated using theprotected LSD test.

In general, leaves exhibited photodynamic injury in both ALA+EN and ALAtreatments just a few hours after exposure to light. This was expressedin the form of wilting and necrosis of leaves. Complete defoliation oftrees was achieved after 30 days following treatment (Table III).

                  TABLE III                                                       ______________________________________                                        Percent Defoliation Of "Golden Delicious"                                     Seedlings In The Greenhouse Over Time                                         Day:     1      3        8    10     17   30                                  Treatment                                                                              % Defoliation                                                        ______________________________________                                        Control.sup.1                                                                          0.77   2.12     2.00 5.00   6.19 6.19                                EN.sup.2 1.13   1.55     3.39 3.39   4.08 4.08                                ALA.sup.3                                                                              27.78  48.61    70.83                                                                              73.61  80.55                                                                              81.25                               ALA + EN.sup.4                                                                         47.27  75.40    94.08                                                                              94.08  96.64                                                                              98.72                               ______________________________________                                         .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate                                                 .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

The experiments indicate that the detachment of the leaves from the stemis caused by the formation of an abscission layer at the leaf petiolelevel. The photodynamic phenomenology expressed by the various applecultivars under greenhouse conditions is briefly described below.

For "Golden Delicious" seedlings, more than 75% of the leaves abscisedby the third day following treatment with ALA+EN and defoliation wascomplete after 30 days (Table III). Except for some negligible browning,EN-treated and control plants exhibited no photodynamic damage (TableIII).

"Red Delicious" seedlings lost 77% of their leaves 17 days aftertreatment with ALA+EN (Table IV). Therefore, the response of "RedDelicious" seedlings to ALA+EN treatment was slower than that observedfor "Golden Delicious". Control and EN-treated "Red Delicious" seedlingsdid not exhibit any noticeable photodynamic damage (Table IV).

                  TABLE IV                                                        ______________________________________                                        Percent Defoliation Of "Red Delicious"                                        Seedlings In The Greenhouse Over Time                                         Day:     1      3        8    10     17   30                                  Treatment                                                                              % Defoliation                                                        ______________________________________                                        Control.sup.1                                                                          0.00   1.65     1.90 1.90   1.90 1.90                                EN.sup.2 0.30   0.79     1.37 1.37   1.37 1.37                                ALA.sup.3                                                                              27.15  41.03    55.30                                                                              56.69  59.47                                                                              60.86                               ALA + EN.sup.4                                                                         21.47  29.70    50.11                                                                              54.06  77.14                                                                              88.83                               ______________________________________                                         .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate                                                 .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

The photodynamic phenomenology of "Winesap"seedlings differed from theprevious two cultivars. Overall, ALA treatment resulted in continuouslyhigher percentage leaf drop over ALA+EN treatment (Table V). Almost 50%defoliation was achieved with ALA alone after the first day of treatment(Table V).

                  TABLE V                                                         ______________________________________                                        Percent Defoliation of "Winesap" Seedlings                                    in The Greenhouse Over Time                                                   Day:     1      3        8    10     17   30                                  Treatment                                                                              % Defoliation                                                        ______________________________________                                        Control.sup.1                                                                          0.30   0.58     1.17 1.17   1.17 1.17                                EN.sup.2 0.89   1.62     2.13 2.13   2.62 2.62                                ALA.sup.3                                                                              49.84  62.74    73.44                                                                              76.71  76.71                                                                              80.97                               ALA + EN4.sup.4                                                                        35.03  45.86    65.27                                                                              65.27  70.15                                                                              73.71                               ______________________________________                                         .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate                                                 .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

                  TABLE VI                                                        ______________________________________                                        Percent Defoliation Of "Prima" Seedlings                                      in The Greenhouse Over Time                                                   Day:     1      3        8    10     17   30                                  Treatment                                                                              % Defoliation                                                        ______________________________________                                        Control.sup.1                                                                          0.79   1.98     2.53 2.53   2.53 2.53                                EN.sup.2 0.79   1.19     1.58 1.58   1.58 1.58                                ALA.sup.3                                                                              37.50  52.78    66.67                                                                              66.67  73.61                                                                              74.31                               ALA + EN.sup.4                                                                         34.65  57.12    61.95                                                                              67.22  75.92                                                                              78.48                               ______________________________________                                         .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate                                                 .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

Again, control and EN treatments exhibited negligible damage (Table VI).

In general, the four apple cultivars exhibited a similar response totreatment with ALA and EN. Indeed, no significant differences amongcultivars were observed in protoporhyrin IX (Proto), MVMg-protoporphyrin monoester (MV MPE), DV Mg-protoporphyrin monoester (DVMPE), MV Protochlorophyllide (MV Pchlide), and DV Protochlorophyllide(DV Pchlide) accumulation in leaves in all treatments (Tables VII-XI).

                  TABLE VII                                                       ______________________________________                                        Leaf Accumulation Of Protoporphyrin IX                                        In Each Of The Four Cultivars Following Treatment                                      Protoporphyrin IX accumulation                                                in nmoles/100 mg protein                                             Cultivar   Control.sup.1                                                                          EN.sup.2                                                                              ALA.sup.3                                                                             ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                          3.16 A.sup.Y                                                                          7.63 A  15.83 A 36.47 B                                   Prima      6.38 A   9.62 A  20.07 A  9.66 A                                   Red Delicious                                                                            5.96 A   5.43 A  15.22 AB                                                                              22.89 B                                   Winesap    4.24 A   7.41 A  24.17 B 25.83 B                                   ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter within a      cultivar are not significantly different.                                     .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

                  TABLE VIII                                                      ______________________________________                                        Leaf Accumulation of Monovinyl Mg-protoporphyrin In                           Monoester Each Of The Four Cultivars Following Treatment                               MV MPE accumulation                                                           in nmoles/100 mg protein                                             Cultivar   Control.sup.1                                                                          EN.sup.2  ALA.sup.3                                                                           ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                          2.32 A.sup.Y                                                                          3.82 A     1.04 A                                                                             5.81 A                                    Prima      2.81 A   4.53 AB   12.11 B                                                                             1.44 A                                    Red Delicious                                                                            2.71 A   2.48 A     1.70 A                                                                             2.27 A                                    Winesap    2.86 A   3.18 A     0.75 A                                                                             0.98 A                                    ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter within a      cultivar are not significantly different.                                     .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

                  TABLE IX                                                        ______________________________________                                        Leaf Accumulation Of Divinyl Mg-protoporphyrin                                Monoester In Each Of The Four Cultivars Following Treatment                            DV MPE accumulation                                                           in nmoles/100 mg protein                                             Cultivar   Control.sup.1                                                                          EN.sup.2                                                                              ALA.sup.3                                                                             ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                          2.57 A.sup.Y                                                                          3.55 A  40.17 B 56.07 B                                   Prima      4.78 A   4.53 A  53.64 B 46.33 B                                   Red Delicious                                                                            2.74 A   2.57 A  45.20 B 102.58 C                                  Winesap    3.03 A   4.44 A  53.76 B 73.34 B                                   ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter within a      cultivar are not significantly different.                                     .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

                  TABLE X                                                         ______________________________________                                        Leaf Accumulation Of Monovinyl Protochlorophyllide                            In Each Of The Four Cultivars Following Treatment                                      MV Protochlorophyllide accumulation                                           in nmoles/100 mg protein                                             Cultivar   Control.sup.1                                                                          EN.sup.2 ALA.sup.3                                                                            ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                         17.71 A.sup.Y                                                                           7.37 A  144.56 B                                                                             220.79 B                                  Prima      20.03 A   8.32 A  257.74 B                                                                             206.82 B                                  Red Delicious                                                                            9.40 A    6.24 A  176.53 B                                                                             403.45 C                                  Winesap    23.64 A  14.12 A  180.20 B                                                                             302.56 C                                  ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter within a      cultivar are not significantly different.                                     .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

                  TABLE XI                                                        ______________________________________                                        Leaf Accumulation Of Divinyl Protochlorophyllide                              In Each Of The Four Cultivars Following Treatment                                      DV Protochlorophyllide accumulation                                           in nmoles/100 mg protein                                             Cultivar   Control.sup.1                                                                          EN.sup.2 ALA.sup.3                                                                            ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                          0.59 A.sup.Y                                                                          0.75 A   2.84 A 6.13 A                                    Prima      1.24 A   0.63 A   2.07 A 4.95 A                                    Red Delicious                                                                            0.62 A   0.24 A   1.74 A 1.13 A                                    Winesap    0.84 A   0.58 A   51.97 B                                                                              8.91 A                                    ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter treatment     within a cultivar are not significantly different.                            .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

However, significant differences among treatments were observed forproto, IX, DV MPE, and MV Pchlide accumulations (Tables VII-XI). Asignificant cultivar x treatment interaction was obtained only in MVPchlide accumulation. As for percent leaf defoliation, no significantdifferences were observed among cultivars. Significant differences were,however, observed among treatments along with a significant cultivar xtreatment interaction (Table XII).

                  TABLE XII                                                       ______________________________________                                        Percent Leaf Defoliation Of Four Apple Cultivars                                       % Leaf Defoliation                                                   Cultivar   Control.sup.1                                                                          EN.sup.2 ALA.sup.3                                                                            ALA + EN.sup.4                            ______________________________________                                        Golden Delicious                                                                          6.19 A.sup.Y                                                                          4.08 A   81.50 B                                                                              98.72 B                                   Prima      2.53 A   1.58 A   74.31 B                                                                              78.48 B                                   Red Delicious                                                                            1.90 A   1.36 A   60.86 B                                                                              88.83 B                                   Winesap    1.17 A   2.62 A   80.97 B                                                                              73.71 B                                   ______________________________________                                         .sup.Y Mean separation for various treatment within a cultivar by LSD at      the 5% level of significance. Means followed by the same letter treatment     within a cultivar are not significantly different.                            .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

Overall, accumulation of proto and extent of leaf defoliation for thecultivars were not significantly different between ALA+EN and ALA;however, they were both significantly higher than EN and controltreatments (Table XIII).

                                      TABLE XIII                                  __________________________________________________________________________    Mean Accumulation Of Tetrapyrroles in nmoles/100 mg                           Protein In "Red Delicious", "Winesap", "Golden                                Delicious" And "Prima" Cultivars And The Percent                              Defoliation For The Four Treatments                                           Proto.sup.1 MV MPE.sup.2                                                                        DV MPE.sup.3                                                                        MV Pide.sup.4                                                                       DV Pide.sup.5                                                                       % Defol.sup.6                             Treatment                                                                           nmoles/100 mg protein                                                   __________________________________________________________________________    Control.sup.7                                                                       4.94 B.sup.Y                                                                        2.68 A                                                                               3.28 C                                                                             17.70 C                                                                             0.82 A                                                                               2.95 B                                   EN.sup.8                                                                             7.52 B                                                                             3.50 A                                                                               3.77 C                                                                              9.01 C                                                                             0.55 A                                                                               2.41 B                                   ALA.sup.9                                                                           18.82 A                                                                             3.90 A                                                                              48.19 B                                                                             189.76 B                                                                            14.65 A                                                                             74.41 A                                   ALA + EN                                                                            23.71 A                                                                             2.62 A                                                                              69.58 A                                                                             283.40 A                                                                            5.28 A                                                                              84.93 A                                   Signif..sup.Z                                                                       *     NS    *     *     NS    *                                         __________________________________________________________________________     .sup.Z Non-significant (NS), significant at the 5% level (*).                 .sup.Y Mean separation within columns by LSD at the 5% level of               significance. Means followed by the same letter for various treatments        within a tetrapyrrole accumulation are not significantly different.           .sup.1 Protoporphyrin IX.                                                     .sup. 2 Monovinyl Mgprotoporphyrin monoester.                                 .sup.3 Divinyl Mgprotoporphyrin monoester.                                    .sup.4 Monovinyl protochlorophyllide.                                         .sup.5 Divinyl protochlorophyllide.                                           .sup.6 Percent defoliation.                                                   .sup.7 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.8 Ethyl nicotinate.                                                      .sup.9 5-aminolevulinic acid.                                            

The accumulation of DV MPE and NV Pchlide was significantly higher inALA+EN treated seedlings than in the other three treatments (TableXIII). However, ALA treated trees also exhibited significantly higher DVMPE and MV Pchlide accumulation in leaves than EN-treated and controlplants. The EN treated seedlings and the controls were not significantlydifferent (Table XIII).

Since the control and the EN treated seedlings did not accumulatesignificant amounts of proto, MVMPE, DV MPE, MV Pchlide, and DV Pchlide,and since almost no defoliation was observed in all four cultivars, itcan be stated that the exogenous ALA and ALA+EN applications weredefinitely responsible for the accumulation of tetrapyrroles and for thesuccessful defoliation of apple trees. When applied with ALA, ENsignificantly increased the dark conversion of exogenous ALA to both DVMPE and MV Pchlide, beyond the level obtained by ALA alone (Table XIII).Therefore, EN can be classified as an enhancer of ALA conversion to MVProtochlorophyllide and to DV MPE.

ALA+EN treatment induced a significantly higher proto accumulation in"Golden Delicious" than in the other treatments. However, ALA+EN treated"Golden Delicious" seedlings did not accumulate higher levels of protothan ALA treated seedlings in other cultivars. Furthermore, in"Winesap", ALA treatment resulted in a significantly higher level ofProto accumulation than that of the Control or EN treatments (TableVII).

ALA-treated seedlings of "Prima" exhibited a higher MVMPE accumulationthan ALA+EN treated plants. However, no significant differences in MVMPE accumulation were exhibited among any of the treatments of "GoldenDelicious", "Red Delicious", and "Winesap" cultivars (Table VIII ) .

"Golden Delicious" , "Prima", and "Winesap" cultivars did not exhibitany significant differences in DV MPE accumulation between the ALA andALA+EN treated plants. However, ALA+EN treated `Red Delicious` treesexhibited a significantly higher DV MPE accumulation than ALA treatedplants (Table IX).

"Red Delicious" and "Winesap" ALA-treated trees exhibited asignificantly higher MV Pchlide accumulation than ALA+EN-treated trees.The other two cultivars did not exhibit a significant difference betweenALA and ALA +EN treatments (Table X).

Only ALA-treated "Winesap" trees exhibited a significant difference intheir DV Pchlide accumulation when compared to the other threetreatments (Table XI).

For all four cultivars, there was no significant difference indefoliation between ALA and ALA+EN treated plants (Table XIII).

Proto, DV MPE, and MV Pchlide accumulation in leaves of the variouscultivars were positively correlated to leaf defoliation (FIGS. 3, 4 and5). In other words, the higher accumulation of proto, DV MPE, and MVPchlide, resulted in an increased rate of defoliation. It was observed,however, that beyond a certain tetrapyrrole concentration level,accumulation of proto, DV MPE and MV Pchlide inhibited defoliation(FIGS. 3, 4 and 5).

Because treated seedlings were not held in cold storage, and because ofthe favorable environmental conditions in the greenhouse, defoliatedseedlings exhibited healthy, green regrowth. This in turn indicates thatshoots and buds were not damaged as a result of these treatments.Interestingly, more regrowth was observed with plants sprayed withALA+EN than with those sprayed with ALA alone. In other words, inaddition to enhanced tetrapyrrole accumulation, the addition of EN toALA appeared to exert growth promoting effects on the defoliated shoots.

Comparison of ALA+EN With Known Chemical Defoliants

Knight's (Knight, J. N., J. Hort. Sci. 58(4):471-476 (1983)) resultsfrom use of FeEDTA and CuEDTA on "Cos's Orange Pippin" apple trees werecomparable to results obtained with the composition according to theinvention. Three percent FeEDTA and 2.1% CuEDTA resulted in 81 and 89%defoliation, respectively, of "Cox's Orange Pippin" apple trees 40 daysfollowing treatment (Knight, J. N. supra), whereas the composition ofALA+EN according to the invention resulted in nearly 99% and 89%defoliation of "Golden Delicious" and "Red Delicious", respectively, 30days following treatment (Table XII). ALA+EN-treated "Prima" trees were78.5% defoliated compared to 74% defoliation of "Winesap" trees (TableXII).

Larsen's (Larsen, F. E., Proc. Internt. Plant Prop. Soc. 17:157-172(1967)) use of 1% S,S,S,-tributylphosphoromtrithionate (DEF) resulted ina complete defoliation of "Winesap" four weeks following treatment andin 80% defoliation of "Rome Beauty" three weeks following treatment.However, DEF did not bring about defoliation of "Red King" appleseedlings (Larsen, F. E., supra). Some excessive damage was reported onthe trees as compared to non-noticeable damage on trees sprayed inaccordance with the invention. Healthy regrowth followed spraying whentrees were kept under favorable environmental conditions.

Defoliation of "Golden Delicious" with Ethrel® at varying concentrationswas significantly inferior to defoliation with the composition accordingto the invention because only 26% leaf drop was obtained (Jones, D. L.,Nichols, D. G., Thompson, W. K., and Jager, L. A., Australian Journal ofExperimental Agriculture and Animal Husbandry 13:460-464 (1973)). Inanother study by Larsen, 8,000 ppm Ethrel resulted in 95% defoliation of"Red Delicious" nursery trees 3-4 weeks following application (Larsen,F. E., J. Amer. Soc. Hort. Sci. 95:662-663 (1970)) .

EXAMPLE 4 Defoliation And Fruit Drop of "Red Delicious" and "GoldenDelicious" Apple Cultivars Under Field Conditions

A total of sixteen apple trees growing at the Pomology Research Farm inUrbana and representing two cultivars, namely "Red Delicious" and"Golden Delicious" were subjected to four different treatments. Eachtreatment was applied to two individual trees. Three branches, 36 inchesin length were selected from each tree for treatment. Therefore, each ofthe four treatments was sprayed on a total of six branches. Fruits werecounted on each branch before spraying.

The four treatments consisted of the following:

(a) control which consisted of solvent only (polyethylene glycol:Methylalcohol:Tween 80:water at 7:2:1:90 (v/v/v/v));

(b) 40 mM ALA; and

(c) 30 mM EN; (d) 40 mM ALA+30 mM EN.

Handling of solutions and sprays was similar to that described inExample 3. Trees of "Red Delicious"were sprayed on 9/15/88 starting at 2p.m. and terminating at 6 p.m. It is important to note that a heavyrainfall occurred in the following morning. Trees of "Golden Delicious"were sprayed on Sep. 16, 1988 starting at 2 p.m. and ending at 6 p.m. Norain showers followed this set of treatments.

To evaluate the degree of photodynamic damage and extent of fruit drop,treated branches were photographed before spraying and on the 10th dayafter spraying for "Golden Delicious" and on the 11th day after sprayingfor the "Red Delicious". Photographic recording was conducted asdescribed earlier, except that a 28-135 mm zoom lens was used. Number offruit drop was recorded every day or every other day and percent fruitdrop was calculated. Data were analyzed in a randomized complete blockdesign.

Treatment of "Red Delicious" branches resulted in neither significantdefoliation nor in fruit abscission (Table XIV), and treated fruits didnot exhibit skin injury. Because "Golden Delicious" and "Red Delicious"responded more or less in the same manner to the application of ALA andALA+EN under greenhouse conditions as shown in Example 3, and becauseboth belong to the same DMV/LDV greening group, the observed differencein the field between the two cultivars is probably not due todifferences in their susceptibility to ALA and ALA+EN treatments, butrather due to the heavy rain that fell the morning after having sprayedthe "Red Delicious" trees.

"Golden Delicious" trees treated with ALA or with ALA+EN underwent a100% defoliation by the 13th day after spraying. By that time, fruitdrop was also 100% (Table XIV). Visual examination of abscised fruitsrevealed the formation of a pronounced abscission layer at the base ofthe pedicel and some fruits exhibited pronounced skin browning. Branchessprayed with EN and solvent only (i.e., control) remained healthy anddid not exhibit a significant level of defoliation or fruit abscissionbeyond normal levels. No injury was observed on sprayed fruit.

Significantly higher leaf defoliation and fruit abscission were observedin ALA and ALA+EN treatments when compared to the control and EN treatedbranches (Table XIV). This was not so in the case of the "Red Delicious"cultivar where no significant difference between treatments was observed(Table XIV).

                  TABLE XIV                                                       ______________________________________                                        Percent Fruit Drop Of "Red Delicious" and "Golden                             Delicious" Cultivars Following Spray Treatments                                           % Fruit Drop                                                      Treatment     Golden Delicious  Red Delicious                                 ______________________________________                                        Control.sup.1 41.67   b.sup.x A.sup.y                                                                         27.94 aA                                      EthylNicotinate.sup.2                                                                       26.27   bA        44.69 aA                                      ALA.sup.3     100.00  aA        41.19 aB                                      ALA + EN      .sup.4 100.00                                                                         aA        30.15 aB                                      ______________________________________                                         .sup.x Mean separation within cultivars by protected LSD at the 1% level      of significance.                                                              .sup.y Mean separation within treatments by protected LSD at the 1% level     of significancce.                                                             .sup.1 Solvent only (polyethylene glycol:methyl alcohol:Tween 80:water at     7:2:1:90 (v/v/v/v)).                                                          .sup.2 30 mM ethyl nicotinate.                                                .sup.3 20 mM 5aminolevulinic acid.                                            .sup.4 20 mM 5aminolevulinic acid + 30 mM ethyl nicotinate.              

Comparison With Other Known Chemical Fruit Harvesters

In other studies (Hartmann, H. T., Fadl, F., and Whisher, J. Calif.Agric. 21(7):5-7 (1967); Unrath, C. R. J. Amer. Soc. Hort. Sci.94:387-391 (1967); Wilson, W. C., and Hendershott, C. H. Proc. Amer.Soc. Hort. Sci. 90:123-129 (1967)), chemicals were used to reduce theforce separating the pedicel to facilitate mechanical harvesting.Therefore, the results from those studies are not comparable to thoseobtained in accordance with the invention where 100% defoliation andfruit drop were achieved following treatment of "Golden Delicious" trees(Table XXIX) without mechanical harvesting.

EXAMPLE 5 Defoliation of Tomato Under Greenhouse Conditions A. MATERIALSAND METHODS

Seedlings of tomato, a DMV/LDV plant species, were grown in a greenhouseunder a fourteen hour lightten hour dark photoperiod in vermiculite fortwenty five days before spraying. (Tomato is accepted in the art as amodel for potato). Two days before spraying, the aging cotyledons wereremoved and the seedlings with old and young developing leaves weresprayed in the afternoon in the greenhouse with 20 mM ALA (about 500g/acre)+15 mM modulator (about 375 g/acre) at a rate of 40 gpa and anaverage droplet size of 75 μm. Eleven different modulators, theidentities of which are shown in Table XXX, were tested. Studies on themode of action of these modulators on cucumber revealed that themodulators acted as enhancers of the conversion of ALA toprotochlorophyllide in cucumber. Depending on the solubility of theparticular modulator, ALA and the modulator were dissolved either in asolvent made up of polyethylene glycol:methanol:Tween 80:water (7:2:1:90v/v/v/v) at a pH of 3.5 or in a solvent made up of polyethylene glycol:Tween 80:ethanol:methanol:water (7:1:45:45:2 v/v/v/v/v) at a pH of 3.5.

The plants were evaluated visually for growth and photodynamic damageover a period of ten days. The plants were periodically photographed torecord response to treatment as described in the previous examples. Theresults of the treatment are shown in Table XV.

                  TABLE XV                                                        ______________________________________                                        Defoliation/Desiccation Of Tomato Seedlings By ALA                            and Nicotinic Acid Or Nicotinamide Modulators                                                % Defoliation/desiccation                                                     Older Leaves                                                                           Younger Leaves                                        ALA    Modulator     Days After Treatment                                     (20 mM)                                                                              (15 mM)       2      7     2     7                                     ______________________________________                                        +      6-aminonicotinamide                                                                         100    100   85    85                                    +      ethyl nicotinate                                                                            95     95    91    91                                    +      2-hydroxynicotinic                                                                          95     95    90    90                                           acid                                                                   +      2-hydroxy-6-methyl                                                                          95     95    74    74                                           pyridine-3-carboxy-                                                           lic acid                                                               +      ethyl 2-methyl                                                                              95     95    71    71                                           nicotinate                                                             +      N-methylnicotin-                                                                            90     90    93    93                                           amide                                                                  +      N-benzyl-N-   90     90    67    65                                           nicotinoyl                                                                    nicotinamide                                                           +      4-hydroxy-7-tri-                                                                            90     90    85    85                                           fluoro methyl-3-                                                              quinoline carboxy-                                                            licacid                                                                +      4-hydroxy-7-methyl-                                                                         89     89    95    95                                           1,8-naphthyridine-                                                            3-carboxylicacid                                                       +      diethyl 3,4-pyridine                                                                        85     85    72    72                                           dicarboxylate                                                          +      niflumic acid 82     82    73    73                                    ______________________________________                                    

An examination of Table XV reveals that tomato leaves were susceptibleto combinations of ALA and nicotinic acid or nicotinamide modulators.Young developing leaves were less susceptible to treatment than theolder, more mature leaves. Overall, ten modulators exhibited 88-90% orbetter defoliation of old leaves. Four of these modulators alsoexhibited 90% or better defoliation of young developing leaves.

EXAMPLE 6 Defoliation Of Cotton Under Greenhouse Conditions

Twenty four day-old vermiculite-grown cotton seedlings were used. Twodays before spraying, aging cotyledons were excised and well-developedprimary leaves (old leaves) as well as developing secondary leaves(young leaves) were treated by spraying with a combination of ALA and agiven modulator. The identities of the modulators tested are set forthin Table XVI. ALA and modulator concentrations were 20 and 15 mM,respectively. Spraying and evaluation of photodynamic defoliation wasexactly as described in Example 5.

The results of the tests are reported in Table XVI.

                                      TABLE XVI                                   __________________________________________________________________________    Defoliation/Desiccation of Cotton By ALA And Various Modulators                                       % DEATH                                               MODULATOR.sup.1         YL.sup.2 T.sup.3 D2.sup.4                                                           T OL.sup.5 D2                                                                      YL T D7.sup.4                                                                       T OL D7                              __________________________________________________________________________    benzyl viologen dichloride monohydrate                                                                87    75   111.sup.6                                                                           111                                  di 2-pyridyl ketoneoxime                                                                              69    81   111   111                                  2,2'-dithiobis(pyridine n-oxide)                                                                      100   100  111   111                                  6,6'-dithiodinicotinic acid                                                                           25    69   111   111                                  methyl5-(benzloxycarbonyl)-2,4-dimethyl-3-                                                            50    57   111   111                                  1-methyl-2-pyrrole carboxaldehyde                                                                     25    37   111   111                                  1-methyl-2-pyrrole carboxylic acid                                                                    31    62   111   111                                  pyrrole-2-carboxaldehyde                                                                              25    87   111   111                                  pyrrole [1,2-a]quinoxaline                                                                            56    100  111   111                                  phenyl 2-pyridyl ketoxime                                                                             79    81   111   111                                  5-chloro-1,10-phenanthroline                                                                          100   100  100   100                                  dimidium bromide        75    94   100   100                                  2,2'-dipyridyl          100   100  100   100                                  ethidium bromide        87    75   100   100                                  methyl viologen dichloride iodide                                                                     100   100  100   100                                  1,10-phenanthroline     100   100  100   100                                  4'-methyl-1,10-phenanthroline                                                                         100   100  100   100                                  1-(carboxymethyl)pyridinium chloride                                                                  94    100  94    100                                  1,1'-diethyl-4,4'-cyanine iodide                                                                      87    100  94    100                                  4,4'-dimethyl-2,2'-dipyridyl                                                                          94    100  94    100                                  1-dodecylpyridinium chloride monohydrate                                                              81    100  94    100                                  1,1'-diethyl-2,2'-cyanine iodide                                                                      87    100  87    100                                  1,1'-diethyl-2,4'-cyanine iodide                                                                      87    100  87    100                                  4,7-dimethyl-1,10-phenanthroline                                                                      87    100  87    100                                  5,6-dimethyl-1,10-phenanthroline                                                                      87    100  87    100                                  5-methyl-1,10-phenanthroline                                                                          87    100  87    100                                  2,2':6'6"-terpyridine   87    100  87    100                                  3,4,7,8-tetramethyl-1,10-phenanthroline                                                               87    100  87    100                                  6-amino nicotinamide    85    100  85    100                                  berberine hydrochloride hydrate                                                                       62    100  75    100                                  5-phenyl-2-(4-pyridyl)oxazole                                                                         75    100  75    100                                  tert-butyl 4-acetyl3,5-dimethyl1-2-pyrrolecarcoxylate                                                 71    100  71    100                                  2-[4-(dimethylamino)styryl]-1-methyl pyridinium iodide                                                71    100  71    100                                  sanguinarine chloride   71    100  71    100                                  1-furfurylpyrrole       69    100  69    100                                  2,4,6-collidine p-toluene sulfonate                                                                   62    100  62    100                                  3-ethyl-2-methyl-4,5,6,7-tetrahydroindol-4-one                                                        62    100  62    100                                  3-amino-2,6-di-methoxypyridine monohydrochloride                                                      50    100  50    100                                  5-amino-2-methoxy pyridine                                                                            50    100  50    100                                  1-ethyl-3-OH-pyridinium bormide                                                                       44    100  44    100                                  6-amino nicotinamide    37    100  37    100                                  ethyl 2-methyl nicotinate                                                                             37    100  37    100                                  ethyl nicotinate        37    100  37    100                                  2-hydroxy-6-methyl pyridine-3-carboxylic acid                                                         37    100  37    100                                  2-hydroxynicotinic acid 25    100  25    100                                  niflumic acid           25    100  25    100                                  dibucaine hydrochloride 14    100  14    100                                  diethyl 3,4-pyridine dicarboxylate                                                                    12    100  12    100                                  ethyl nicotinate        91    95   91    95                                   2-hydroxy nicotinic acid                                                                              90    95   90    95                                   2-hydroxy-6-methyl pyridine-3-carboxylic acid                                                         74    95   74    95                                   ethyl 2-methylnicotinate                                                                              71    95   71    95                                   5-nitro-1,10-phenanthroline                                                                           100   94   100   94                                   phenanthridine          81    94   81    94                                   ethyl 3,5-dimethyl-2-pyrrolecarboxylate                                                               75    94   75    94                                   2-[4-(dimethylamino)styryl]-1-ethyl pyridinium iodide                                                 62    87   75    94                                   isocarbostyril          75    94   75    94                                   2,3-dihydroxypyridine   62    94   62    94                                   2-chloro-6-methoxypyridine                                                                            50    94   50    94                                   4-hydroxy-7-trifluoro methyl-3-quinoline carboxylic                                                   50    94   50    94                                   1-(dimethylamino)pyrrole                                                                              44    94   44    94                                   4-hydroxy-7-methyl-1,8-naphthridine-3-carboxylic acid                                                 43    93   43    93                                   N-methylnicotinamide    93    90   93    90                                   4-hydroxy-7-trifluoro methyl-3-quinoline carboxylic                                                   85    90   85    90                                   N-benzyl-N-nicotinoylnicotinateamide                                                                  65    90   65    90                                   4-hydroxy-7-methyl-1,8-naphthridine-3-carboxylic acid                                                 95    89   95    89                                   citrazinic acid         81    87   81    87                                   propidium iodide hydrate                                                                              75    87   75    87                                   2-hydroxy-4-methyl quinoline                                                                          71    87   71    87                                   3-cyano-4,6-dimethyl-2-hydrochloride                                                                  62    87   69    87                                   N-benzyl-N-nicotinoyl nicotinatemide                                                                  56    87   56    87                                   diethyl-2,4-dimethyl pyrrole-3,5-dicarboxylate                                                        56    87   56    87                                   4-(dimethylamino)pyridinium bromide perbromide                                                        56    87   56    87                                   1-(2-cyanoethyl)pyrrole 37    81   50    87                                   N-methylnicotinamide    25    87   25    87                                   diethyl3,4-pyridine dicarboxylate                                                                     72    85   72    85                                   niflumic acid           73    82   73    82                                   4,7-diphenyl-1,10-phenanthroline                                                                      75    81   75    81                                   8-hydroxy-5-nitroquinoline                                                                            75    81   75    81                                   5,7-dichloro-8-hydroxyquinoline                                                                       81    75   81    75                                   2,6-dimethoxypyridine   50    75   50    75                                   5-chloro-8-hydroxy-7-iodoquinoline                                                                    44    69   44    75                                   5,7-dibromo-8-hydroxyquinoline                                                                        44    75   44    75                                   bis-N-methyl acridinium nitrate                                                                       31    75   31    75                                   2,9-dimethyl-4,7-phenyl-1,10-phenanthroline                                                           25    25   25    25                                   3-hydroxypicolinic acid 12    19   19    19                                   1-isoquinolinecarboxylic acid                                                                         12    12   12    12                                   picolinic acid          12    12   12    12                                   __________________________________________________________________________     .sup.1 ALA and modulator concentrations were 20 and 15 mM respectively.       .sup.2 YL = young leaves.                                                     .sup.3 T = treated.                                                           .sup.4 D2, D7 = damage 2 and 7 days after spraying.                           .sup.5 OL = old leaves.                                                       .sup.6 The number 111 in each column refers to the development of             nutrition deficiency symptoms due to an error in the Hoagland solution.       The plants died from nutritional toxicity befor the ten day photodynamic      injury was evaluated.                                                    

Based on the photodynamic injury observed on day two, compositionscontaining ALA and modulators 488 and 814, respectively, were effectivedefoliants. Overall, the older mature leaves were more susceptible thanthe younger developing leaves, as was observed with tomato. Fifty sevenof the modulators tested exhibited 90-100% defoliation of older leaveswhile twelve modulators exhibited similar performance on younger leaves.Seven of the modulators exhibited 100% defoliation of both young and oldleaves.

These examples serve to demonstrate the novel concept of the presentinvention. The photodynamic mode of action is different from other knownmodes of action in two main respects: (a) it is dependent on thebiosynthesis and accumulation of tetrapyrroles in the foliage of livingplants; and (b) the accumulated tetrapyrroles render the foliage of theplants light-sensitive so that upon subsequent exposure to light, a verydamaging photodynamic effect is produced which results in desiccation ofthe foliage without death of the plants.

δ-Aminolevulinic acid is a natural metabolite present in all livingcells; it is a natural component of the biosphere and is readilybiodegradable. The same is true for the products of ALA dark-metabolism,i.e., the tetrapyrrole intermediates of the Chl biosynthetic pathway,which have been demonstrated to disappear very rapidly upon exposure ofthe plant to light. Similarly, modulators which are naturally occurringvitamins or derivatives thereof, e.g., ethyl nicotinate, are expected tobe readily biodegradable and to have no adverse impact on theenvironment. It therefore appears that the photodynamic desiccatingcompositions and methods of the present invention employing ALA and/orvitamins or derivatives thereof are likely to have no adverse impact onthe environment.

Further examples of compositions and applications within the spirit andscope of this invention are described in copending application Ser. No.895,529 and in Rebeiz, C. A. et al., CRC Critical Reviews in PlantSciences, 6(4):385-436 (1988).

I claim:
 1. A method for defoliating a deciduous tree comprising thesteps of:(a) contacting said tree with a defoliating effective amount ofδ-aminolevulinic acid; and (b) allowing said contacted tree of step (a)to be exposed to light.
 2. A method as recited in claim 1, wherein saidcontacted tree of step (a) is exposed to a substantial absence of lightat wavelengths of 300 to 700 mM before being exposed to light in step(b).
 3. A method as recited in claim 1, wherein said contacted tree ofstep (a) is exposed to said substantial absence of light for about 1 to8 hours before being exposed to light in step (b).
 4. A method asrecited in claim 1, wherein said tree is exposed to light in step (b)for a period of time sufficient to oxidize most unsaturated membranelipoproteins of the foliage of said plant.
 5. A method as recited inclaim 1, wherein said tree is exposed to light in step (b) in the formof natural daylight for a period of about 1 to 14 days.
 6. A method forcausing defoliation and fruit drop in a deciduous fruit tree comprisingthe steps of:(a) contacting said tree with an amount effective to causedefoliation and fruit drop in said tree of δ-aminolevulinic acid; and(b) allowing said contacted tree of step (a) to be exposed to light. 7.A method as recited in claim 6, wherein said contacted tree of step (a)is exposed to a substantial absence of light at wavelengths of 300 to700 mM before being exposed to light in step (b).
 8. A method as recitedin claim 6, wherein said contacted tree of step (a) is exposed to saidsubstantial absence of light for about 1 to 8 hours before being exposedto light in step (b).
 9. A method as recited in claim 6, wherein saidtree is exposed to light in step (b) for a period of time sufficient tooxidize most unsaturated membrane lipoproteins of the foliage of saidplant.
 10. A method as recited in claim 6, wherein said tree is exposedto light in step (b) in the form of natural daylight for a period ofabout 1 to 14 days.
 11. A method as recited in claim 1, wherein saideffective amount of δ-aminolevulinic acid is at least 15 mM.
 12. Amethod as recited in claim 6, wherein said effective amount ofδ-aminolevulinic acid is at least 15 mM.